Oral tolerance is a long held observation that oral antigen results in systemic hypo-responsiveness. Recently, there is increased interest in oral tolerance following the demonstration that orally administered antigen can suppress several animal models of autoimmune disease. In addition, the oral administration of autoantigen is being tested in human autoimmune diseases and in transplantation. Nonetheless, a great deal remains to be learned about the basic mechanisms that underlie immune responses in the gut associated lymphoid tissues (GALT). This section of the program project grant will investigate the role of antigen affinity and cytokines in oral tolerance in the EAE model. Effector mechanisms of immune tolerance induced by oral antigen have been defined and related to dose of protein fed. Low doses of protein favor induction of regulator T cells that secrete Th2 cytokines and TGF-beta whereas higher doses induce anergy or deletion. The nature of the antigen and its interaction with the GALT has not been well studied. In addition, an understanding of the importance of Th2 cytokines in regulating oral tolerance and their use as enhancers has not been studied. Finally, we have described a new subset of T cells termed Th3 cells that are associated with oral tolerance and the nature of these cells has not been well characterized. We propose to study the factors that promote low affinity interactions between peptide-MHC-TCR leading to potential generation of TGF-beta and Th2 responses. In our investigations, we will study altered peptides (APLs), which have been generated by altering TCR or MHC contact residues of the encephalitogenic MPB and PLP peptides; these APLs will provide unique tools to address the issue of strength of signal in the induction of oral tolerance. We also have available MBP and PLP TcR transgenic mice to undertake these studies. Thus the three specific aims of our proposal are: 1) What is the effect of native, altered and cross-reactive peptides on oral tolerance? 2) What are the mechanisms associated with the induction of TGFbeta secreting (Th3 type) cells? And 3) What are the mechanisms associated with enhancement of oral tolerance by orally administered cytokines? Addressing these questions will provide basic information regarding mechanisms of the induction of oral tolerance and immune responses in the gut associated lymphoid tissue that will have implications for the use of oral antigen to treat autoimmune diseases such as multiple sclerosis.